ABSTRACT/PROJECT SUMMARY Despite clinical advances, the fact remains that humans cannot regenerate injured tissue. Instead, tissue trauma stimulates inflammation, which in turn induces fibrosis and ultimately the formation of scar tissue. Trauma-induced inflammation is primarily regulated by macrophages; key regulators of fibrosis. Although current therapies to control human fibrotic diseases include agents that target macrophages, new data indicate these treatments negatively impact regenerative ability. This outcome results from our lack of knowledge about how macrophages regulate endogenous tissue regeneration. A mammalian model of tissue regeneration is necessary to directly study how macrophages regulate the injury response to fill this knowledge gap. The discovery that African spiny mice (Acomys) can regenerate complex tissues of the external ear including skin, hair follicles, nerves, muscle and cartilage provides such a model. Furthermore, because laboratory mice produce scar tissue in response to an identical injury, this comparative system provides a unique opportunity to directly study how macrophage phenotypes regulate regeneration and scarring. The long-term goal is to develop clinical interventions that stimulate regeneration and inhibit fibrosis. The objective of this proposal is to identify macrophage subtypes that regulate regeneration and manipulate inflammation to stimulate regeneration in response to injury. This proposal tests the central hypothesis that specific macrophage subtypes stimulate a regenerative extracellular environment and that phenotype switching from one state to another is critical to the outcome of injury. This hypothesis is based upon published and preliminary studies indicating general macrophage phenotypes are associated with regeneration, that Acomys and Mus exhibit different cytokine and inflammatory gene profiles and that production of a regenerative or scarring extracellular environment corresponds to specific macrophage phenotypes observed in Acomys or Mus. Guided by strong preliminary data, this hypothesis will be tested by pursuing the following two specific aims: (1) identify specific macrophage phenotypes that regulate tissue regeneration and scarring in Acomys and Mus. Parallel in vivo and in vitro studies will integrate the timing and extracellular influence of macrophage subtypes and macrophage origin using newly developed regeneration assays; and (2) test the role of macrophage-produced arginase on tissue regeneration through the use of a newly identified immunomodulatory agent and genetically engineered mice with macrophages that lack arginase. The approach is innovative, in the applicant's opinion, because it departs from the status quo of investigating wound healing, and instead, focuses on understanding endogenous tissue regeneration in a unique model of mammalian tissue regeneration. The proposed research is significant because it is expected to provide new and fundamental insight into how macrophages coordinate the inflammatory and immune reaction to stimulate a regenerative response to injury.